CLOSTRIDIUM PASTEUEIANUM. 353 



-are stained a deep blue-black by iodine. The ripe spore escapes 

 through the wall of the mother-cell in a longitudinal direction. 

 The great resemblance between this clostridium and the butyric 

 acid bacteria described in a previous section is not only morpho- 

 logical, but also extends to the fermentative capacity. For example, 

 Clostridium Pasteurianum acts on sugar in such a manner that 

 both volatile acids : butyric acid and acetic acid (4:1), and gases : 

 carbon dioxide and hydrogen (60-75 P er cent - f H. by volume) 

 .are formed. The importance to us of this fission fungus on the 

 .present occasion consists in its behaviour towards nitrogen, which 

 gas it absorbs from the atmosphere, fixes it and employs it in the 

 elaboration of organic substances. The energy necessary thereto 

 is supplied and liberated by the decomposition of sugar; conse- 

 quently it is easy to understand that a definite relation exists 

 between the amounts of sugar fermented and of nitrogen com- 

 bined. This ratio was determined by Winogradsky as 2.5-3 

 m.grms. of nitrogen to 1000 m.grms. of dextrose. This element 

 (N.) when in a state of combination is not only valueless to 

 Glostridium Pasteurianum^ but when present in large quantity 

 even injurious thereto. For the cultivation of the microbe 

 which cannot be carried out on the ordinary nutrient media 

 (gelatin, bouillon) in use, though it grows on sliced potatoes 

 use is made, preferably, of an aqueous solution containing i gram 

 K 3 P0 4 , 0.5 gram MgS0 4 , 0.01-0.02 gram NaCl,FeS0 4 ,MnS0 4 , 

 a little CaC0 3 (for fixing the acids), and 20-40 grams of dextrose 

 per litre. Glostridium Pasteurianum is strictly anaerobic, and in 

 the soil is therefore obliged to rely on the co-operation of aerobic 

 fission fungi, which remove the injurious oxygen from its sphere 

 of influence, surround it with an atmosphere of nitrogen, and as 

 a reward for this service have the opportunity of consuming the 

 nitrogenous substances elaborated and excreted by the Glostridium. 

 Winogradsky observed two species of such assistant organisms, 

 detailed mention of which can, however, be omitted, it being 

 sufficient for our purpose to have referred to this new case of 

 symbiosis. 



Whether the just-named faculty of fixing nitrogen is also pos- 

 sessed by the organisms akin to Clostridium Pasteurianum, e.g. 

 Prazmowski's Glostridium butyricum, still remains undetermined. 

 WINOGRADSKY (III.) examined fifteen species of soil bacteria in 

 this connection, with only negative results. In the higher fungi 

 K. PURIEWITSCH (I.), in an improved continuation of Berthelot's 

 researches, showed that both Aspergillus niger and Penicillium 

 glaucum fix free nitrogen. Their potency is, however, but slight, 

 and is not to be compared with that of Clostridium Pasteurianum, 

 since it is not manifested in media devoid of nitrogen. According 

 to H. JUMELLE (I.), Spirillum luteum, also, is capable of thriving 

 in media free from nitrogen. 



The proof of the fact that the fixation of free nitrogen occurs 

 VOL. i. z 



